Characterization of Alzheimer's Mutations in ADAM10.

ADAM10 中阿尔茨海默病突变的表征。

• 批准号: 8890722
• 负责人: RUDOLPH Emile TANZI
• 金额: $ 33.54万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-30 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8890722
• 关键词: Age of Onset; Alzheimer' s Disease; Amyloid; Amyloid beta-Protein Precursor; Animal Model; Attenuated; Biochemical; Biological Assay; Biotinylation; Brain; Catalytic Domain; Cell membrane; Cerebrum; Cognitive; Collection; Data; Density Gradient Centrifugation; Development; Disease; Dominant-Negative Mutation; Electrophysiology (science); Etiology; Exhibits; Family; Foundations; Gene Expression; Genes; Genetic; Genotype; In Vitro; Knock-in Mouse; Late Onset Alzheimer Disease; Linkage Disequilibrium; Long-Term Potentiation; Medical Genetics; Memory; Metalloproteases; Missense Mutation; Molecular; Molecular Chaperones; Mus; Mutation; National Institute of Mental Health; Pathogenesis; Pathogenicity; Peptide Hydrolases; Phenotype; Play; Prevention; Production; Property; Proteins; Regulation; Reporting; Role; Sampling; Senile Plaques; Single Nucleotide Polymorphism; Staining method; Stains; Structure; Surface; System; Testing; Tg2576; Transgenic Mice; Transgenic Organisms; Validation; Variant; Yeasts; amyloid precursor protein processing; base; cognitive function; disease phenotype; enzyme activity; familial Alzheimer disease; genome wide association study; in vivo; mouse model; mutant; neuropathology; novel; overexpression; proband; protein folding; secretase; segregation; spatial memory; targeted treatment; trafficking

DESCRIPTION (provided by applicant): Abundant clinical, genetic, and biochemical data support the hypothesis that abnormal processing of the amyloid precursor protein (APP) and the cerebral accumulation of its metabolite, A�, play key roles in the etiology and pathogenesis of Alzheimer's disease (AD). A� is generated via serial cleavage of APP by �- and �- secretase. In contrast, cleavage of APP by �-secretase precludes A� production; the major �-secretase in the brain is ADAM10. We recently reported two rare missense mutations in ADAM10 that strongly co- segregates with late-onset AD (LOAD). Both are prodomain mutations that were found in 7 of 1000 NIMH and NIA LOAD families tested (average age of onset, ~70 years). We have also previously reported that both mutations significantly attenuated �-secretase activity and elevated A� levels in vitro. To further validate and expand upon these novel findings, we propose to 1. Search for additional novel familial LOAD mutations in ADAM10; 2. Validate the pathogenicity of these two missense mutations in vivo; and 3. Determine the molecular mechanism by which these two mutations impair �-secretase activity. For Aim 1, we will search for additional novel familial LOAD mutations in ADAM10, through targeted re-sequencing (and genotyping) of 2454 additional AD families (N=6516 subjects) from the NIMH and NIA AD family collections. Our family-based GWAS on >900 NIMH and NIA AD families suggest the existence of additional AD-associated SNPs in ADAM10. With regard to Aim 2, we will attempt to validate the pathogenicity of these two mutations in vivo in transgenic mice (already generated) overexpressing either wild-type (WT) or mutant (Q170H, R181G, artificial dominant-negative) forms of ADAM10. These mice have already been crossed with Tg2576 (APPswe) AD mice so that we can also test for effects of these mutations on neuropathological and cognitive phenotypes of AD. Our preliminary in vivo results reveal that the ADAM10 prodomain mutations attenuate non-amyloidogenic processing of APP, and that senile plaque counts and A� levels were significantly increased in the brains of APPswe/ADAM10 double transgenic mice expressing mutant forms of ADAM10, as compared to those expressing WT forms. Moreover, to test the impact of the prodomain mutations under more physiologically relevant conditions, we have added a plan to generate and characterize ADAM10 mutant knock-in mice. Finally, in Aim 3, we will investigate the molecular mechanism by which the prodomain mutations down- regulate ADAM10 �-secretase activity. Briefly, we will test for effects of these mutations on three prodomain functions: regulation of enzyme activity, intracellular trafficking, and intramolecular chaperoning. At the completion of this project, we hope to provide genetic, biochemical, and mechanistic evidence validating the pathogenicity of late-onset familial AD mutations in ADAM10. Moreover, the data emerging from the proposed study would serve as a firm foundation for the discovery and development of new therapies targeting ADAM10 for the treatment and prevention of this devastating disease.

描述（由适用提供）：丰富的临床，遗传和生化数据支持以下假设：淀粉样蛋白前体蛋白（APP）的异常加工以及其代谢物的大脑积累，A'，A'，A'，在阿尔茨海默氏病（AD）的病因和病原体中起关键作用。通过``App and secretase and secravase of app cleavage of app cleavage of App clavage of off）生成。相比之下，通过 - 分泌酶对APP的裂解排除了生产；大脑中的主要 - 分泌酶是ADAM10。我们最近报道了ADAM10中有两个罕见的错义突变，这些突变与晚期AD（负载）强烈共聚。两者都是prodomain突变，在1000名NIMH和NIA负载家族中的7个（平均发病年龄，约70年）中发现。我们先前还报告说，这两个突变都显着减弱 - 分泌酶活性，并在体外升高了水平。进一步验证并扩大了这些新发现的结果，我们建议1。在ADAM10中寻找其他新型的农场负荷突变； 2。验证体内这两个错义突变的致病性；和3。确定这两个突变会损害 - 分泌酶活性的分子机制。对于AIM 1，我们将通过从NIMH和NIA AD Family Collections的2454个其他AD家族（n = 6516名受试者）进行针对性的重新测量（和基因分型）来搜索ADAM10中的其他新型农业负荷突变。我们基于家庭的GWA在> 900 NIMH和NIA AD系列上表明ADAM10中存在其他与广告相关的SNP。关于AIM 2，我们将尝试验证转基因小鼠（已经产生的）过表达野生型（WT）或突变体（Q170H，R181G，人造主导性）ADAM10中的这两个突变的致病性。这些小鼠已经与TG2576（AppSwe）AD小鼠杂交，因此我们还可以测试这些突变对AD神经病理学和认知表型的影响。我们的初步体内结果表明，ADAM10 Prodomain突变减弱了APP的非淀粉样蛋白原的处理，并且与表达ADAM10的表达ADAM10相比，Appswe/Adam10双转基因小鼠的大脑中的老年斑块和A水平显着升高，与表达WT形式相比。此外，为了测试在更加相关的条件下Prodomain突变的影响，我们添加了一个计划，以生成和表征ADAM10突变体敲门小鼠。最后，在AIM 3中，我们将研究Prodomain突变下调ADAM10-分泌酶活性的分子机制。简而言之，我们将测试这些突变对三个的影响 Prodomain功能：调节酶活性，细胞内运输和分子内伴侣。该项目完成后，我们希望提供遗传，生化和机械证据，以验证ADAM10中晚期家庭AD突变的致病性。此外，拟议的研究中出现的数据将成为发现和开发针对ADAM10的新疗法的坚定基础，以治疗和预防这种毁灭性疾病。